It has been known that heparin-binding proteins, among all, those proteins classified into the fibroblast growth factor (hereinafter, referred to as “FGF”) family strongly bind to heparin and heparan sulfate (sulfated polysaccharides) by a non-covalent bonding mode. It has been also known that when a heparin-binding protein such as fibroblast growth factor is mixed with a sulfated polysaccharide such as heparin, the biological activity and physical properties of the heparin-binding protein are altered to change its function; sometimes, such a heparin-binding protein may acquire higher function. However, even if a sulfated polysaccharide was mixed with, the expected augmentation of the protein function has been limited. Besides, when such a mixture is used as a pharmaceutical composition, undesirable physiological activity attributable to free sulfated polysaccharides has caused some problems.
For the purpose of augmentation of function of heparin-binding proteins, preparation of a heparin-binding protein to which heparan sulfate(s) is(are) selectively conjugated by covalent bond was intended, and a method of producing such a heparin-binding protein was invented. However, according to that invention, the sulfated glycosaminoglycan sugar chains covalently bonded to the heparin-binding protein were a mixture of heparan sulfate and chondroitin sulfate. Since the results of analysis have revealed that augmentation of function of heparin-binding proteins is attributable to heparan sulfate alone, a method of preparing a functionalized heparin-binding protein has been desired by which heparan sulfate is preferentially added and little chondroitin sulfate is added.    Patent document 1: “HEPARIN-BINDING PROTEINS MODIFIED WITH SUGAR CHAINS, METHOD OF PRODUCING THE SAME AND PHARMACEUTICAL COMPOSITIONS CONTAINING SAME”, Japanese Patent No. 3318602 (2002), Imamura T, Asada M, Oka S, Suzuki M, Yoneda A, Ota K, Oda Y, Miyakawa K, Orikasa N, Matsuda C, and Kojima T.    Non-patent document 1: Yoneda A, Asada M, Oda Y, Suzuki M, and Imamura T. “Engineering of an FGF-Proteoglycan Fusion Protein with Heparin-Independent, Degradation-Augmented, Mitogenic Activity.” Nature Biotechnology 18 (6), 641-644 (2000)    Non-patent document 2: Yoneda A, Asada M, and Imamura T, “Modification of the Activity of Heparin-Binding Growth Factor FGF-1 by Fusion with Syndecan [in Japanese]”, Saiboukougaku 19 (9), 1338-1340 (2000)    Non-patent document 3: Asada M, Yoneda A, Imamura T. “Engineering of a Heparin-Binding Growth Factor with Heparan Sulfate Sugar Chains”, Trends in Glycoscience and Glycotechnology 13 (72) 385-394 (2001)